Description du sujet de thèse

Nonlinear optical microscopy makes it possible to study biological tissues in 3D over depths of a few hundreds of micrometers with subcellular resolution. LOB is pioneering the use of label-free nonlinear optical signals to study the structure and evolution of biological tissues (embryo, skin, brain) with sub-cellular resolution. In particular, fluorescence lifetime imaging (FLIM) provides information on the metabolic state of cells (see image) and polarized third-harmonic generation microscopy (THG) reveals high-index structures such as myelinated axons. The goal of this project is to develop an efficient imaging approach for quantifying cell metabolism and myelin distribution in nervous tissues.
During the first year of the project, the work will focus on two aspects. First, the two contrast modalities will be adapted for imaging immobilized zebrafish larvae. This will require to optimize an existing lab-built microscope equipped with a tunable femtosecond laser source, time-resolved detection for FLIM, and polarization control. High-resolution FLIM-THG images will be recorded in the spinal cord region. Second, the PhD student will develop myelin and metabolic scores based on these images. For the following two years, this work will include a methodological and an applicative part. On the methodological side, the aim is to pursue the development of FLIM-THG imaging. This will include the development of polarization and beam shaping methods for enhancing the sensitivity of THG to myelinated axons, and the acceleration of FLIM imaging with improved detectors. On the application side, these developments will be used (i) to better understand lipid and metabolic disorders using zebrafish embryos as a live model, and (ii) to record large-area maps of myelin distribution in ex vivo healthy and pathological tissue samples. Image analysis methods will be implemented to extract scores at scales ranging from sub-micrometer to centimeter.

Contexte de travail

The project will take place in the 'Advanced microscopies' pole of the Lab for Optics and Biosciences at Ecole Polytechnique (LOB). The work will involve daily interactions with a group of 3-4 people, within a microscopy team of ~25 persons and in collaboration with the Paris Brain & Spine Institute. The project involves experimental optics, data analysis, numerical simulations, and biological samples manipulation.

Contraintes et risques

Femtoseconde laser